Sensitivity of Steroid Enzyme Immunoassays. : Use of 3,3', 5,5'-Tetramethylbenzidine as a Chromogen in the Assay System with Glucose Oxidase Label

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The use of 3,3', 5,5'-tetramethylbenzidine (TMB) as a chromogen in a testosterone enzyme immunoassay system with a glucose oxidase label is described. Glucose oxidase activity was measured with glucose, peroxidase and TMB in acetate-citric acid buffers ; the optimum pH value was found to be 4.2. The glucose oxidase-labeled antigen was prepared by the N-succinimidyl ester method. In the competitive enzyme immunoassay, the bound and free enzyme-labeled antigens were separated by a double antibody method. A dose-response curve with a high sensitivity was obtained by the use of a minimum amount of the label at an appropriate dilution of anti-testosterone antiserum (K_a=2×10^<10>M^<-1>). The amount of testosterone needed to displace 50% of the bound lab el was 24 pg. The sensitivity was higher than that of the assay system using 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid), ABTS.
公益社団法人日本薬学会の論文
1988-05-25